Grantee Research Project Results
Final Report: Inhalability of Particulate Matter in Laboratory Animals
EPA Grant Number: R827996Title: Inhalability of Particulate Matter in Laboratory Animals
Investigators: Asgharian, Bahman
Institution: The Hamner Institutes
EPA Project Officer: Chung, Serena
Project Period: January 17, 2000 through January 16, 2002
Project Amount: $335,903
RFA: Airborne Particulate Matter Health Effects (1999) RFA Text | Recipients Lists
Research Category: Air Quality and Air Toxics , Particulate Matter , Air , Human Health
Objective:
The overall objective of the research project was to develop improved animal dosimetry models for use in risk assessments of particulate matter (PM). Accurate risk assessments of PM require a thorough understanding of the relationships between ambient exposure, dose to the respiratory tract, and responses. A number of studies have shown that acute exposure to PM is related to increased mortality and morbidity in humans. Since similar findings have been difficult to observe in animals, the U.S. Environmental Protection Agency (EPA), along with several other organizations, have developed major research initiatives that primarily focus on compromised animal models. To make the most of this new data, quantification or estimation of the particulate dose responsible for given effects must be determined. Extrapolation of animal deposition study results to human exposure scenarios requires comparable, but not identical, exposure environments. Not all particles in the air are inhalable by rats, even though they may be in the inhalable range for humans. Deposition in the upper respiratory tract (URT) significantly affects the potential for particles to deposit in the lower respiratory tract (LRT). Therefore, determining the probability that rats will inhale particles is critical for accurate risk assessments. Uncertainty with regard to dose to the lungs of animals heavily is influenced by the lack of good models of deposition. Our primary research objectives were to: (1) expose female Long-Evans rats to fine and coarse aerosols, measure nasal and lung deposition of particles, and use the information to calculate particle inhalability fraction; and (2) use this data to develop and refine particle dosimetry models.
Summary/Accomplishments (Outputs/Outcomes):
To determine regional lung deposition of inhaled particles, we exposed female Long-Evans rats to monodisperse particles in several ways: the exposure atmosphere passing directly through the nose of the animal, head-only exposure, nose-only exposure, and whole-body exposure. In method one, anesthetized rats were exposed to particles while their breathing was controlled externally, allowing deposition in the nasal cavity only. The flow was directed through the nose of the animal so that the particle concentration in the exposure air was the same as that entering the nose of the animal. Head deposition efficiency was measured for inhalation and exhalation. The deposition efficiency data was fitted to a function that depended on the impaction parameter. The head deposition fraction efficiencies could directly be inserted into the respiratory tract deposition model for more accurate assessment of lung deposition. In methods two-four, rats briefly were exposed to monodisperse polystyrene latex particles (Spherotech Inc., Libertyville, IL) via head-only, nose-only, and whole-body exposures while their breathing parameters were being measured. This procedure allowed deposition in the nasal region and LRT under laboratory exposure scenarios. Animals breathed from the air stream so that the inhaled concentration of particles was different from that in the exposure air. Nasal and lobar deposition of particles was measured, and deposition fractions in various regions of the respiratory tract were obtained.
In the case of 100 percent inhalability, the deposition characteristics of large, inhalable particles in rat nasal passages were investigated by determining deposition efficiencies of these particles in a nasal mold of an F344 rat for steady-state and pulsating flow conditions. Particles with geometric diameters ranging from 0.9 to 4 µm, and flow rates ranging from 300 to 900 ml/min, were employed for simulated inspiratory and expiratory flow situations. An optically clear acrylic mold was fabricated from a life-size metal cast that comprised the nares, nasal cavity, pharynx, and larynx. Deposition efficiencies were measured for each flow situation and plotted as functions of particle inertia.
Inspiratory and expiratory deposition efficiencies were similar for a given flow condition. Deposition efficiencies for pulsating flow markedly were higher than those for steady flows. The results for pulsating flows indicated higher deposition efficiencies than were found in previous studies in the literature performed with live rats. These differences may be due to uncertainties in particle inhalability, clearance, and flow rate in the previous studies, as well as differences between the nasal geometries of live rats and the geometry of the nasal mold made from a postmortem cast. The results suggest that the pulsating nature of breathing is an important consideration when determining the deposition of fine and coarse particles.
In the next series of experiments, streams of fine and coarse aerosols were passed through the nasal airways of female Long-Evans rats to determine the entire range of impaction-related aerosol deposition efficiencies. Aerosols traveled through the nasal airways in steady-state and pulsating flows for simulated inspiratory and expiratory scenarios. The nasal region was isolated from the rest of the respiratory tract, and the flow volume and pulsating rate through the nasal region were controlled externally. Polystyrene latex microspheres with geometric diameters ranging from 0.9 to 4 µm and average flow rates ranging from 220 to 639 ml/min were used.
Deposition rose sharply with increased particle inertia for all exposure scenarios. Expiratory deposition efficiency appeared to be somewhat higher than inspiratory deposition efficiency for both steady-state and pulsating flow conditions. Pulsating flows yielded significantly higher deposition than steady-state flows. This result indicated the importance of considering fluid accelerations inherent in normal breathing when determining aerosol deposition dominated by inertial impaction. Slightly higher results were found for the case of pulsating flow in the present study compared with the previous in vivo deposition studies. Variability in the data, which was suspected to primarily result from the difficulty in surgical procedures, was in excess of expected intersubject variability.
Comparison of the results of nasal mold and anesthetized rat experiments revealed that deposition efficiencies in the nasal mold were similar to those in the live anesthetized rat for inspiratory and expiratory flows under steady-state and pulsating breathing conditions. Thus rat nasal molds may well be suitable surrogates for studies on the deposition of large particles using live rats.
A condensation monodisperse aerosol generator (CMAG) (TSI, St. Paul, MN) was used to produce aerosol particles for inhalation exposures. Monodisperse aerosols were generated from a solution of radiolabeled iron chloride (59FeCl3). Long-Evans rats were exposed to the radiolabeled particles to determine nasal, lung lobar, and total lung deposition fractions. The inhalable fraction of particles in the exposure air varied with the type of exposure chamber. Animals were exposed to airborne particles via single-chamber plethysmographs (Buxco Electronics, Sharon, CT), head-only, or whole-body, and Cannon exposure chamber nose-only. Particle inhalability fraction in the Cannon nose-only exposure scenario also was obtained by comparing the measured deposition fractions with the predicted values for 100 percent inhalability.
Monodisperse aerosol particles with aerodynamic diameters ranging from 0.9 to 4 µm were generated by the CMAG. The generated aerosol was diluted with clean air to reach the target exposure concentration. In the first series of experiments, the exposure air was led to a six port manifold that distributed the exposure atmosphere evenly among the chambers housing the rats (either head-only or whole-body Buxco animal tubes). Five rats in individual chambers were exposed per exposure event.
The Buxco animal holding tubes enabled measurement of breathing parameters during exposure. An aerosol particle sizer (APS) (Model 3320, TSI Inc., St. Paul, MN) monitored particle concentration and aerodynamic diameter during exposure. A filter sample was placed at one port of the manifold to collect aerosols from the exposure line. A constant flow through the filter sample was maintained using house vacuum and a mass flow controller (MKS Instruments, Model 246B, Andover, MA). The filter sample was analyzed by scintillation counting (Packard Cobra II Series, Packard Instrument Company, Downers Grove, IL) to determine the total amount of exposure activity.
Animals were euthanized immediately following exposure. The nasal passages, larynx, esophagus, stomach, duodenum, trachea, and lung lobes were dissected and counted by scintillation to determine the amount of deposited activity. Deposition fractions in the various tissues were determined as follows,
(1)
where DFi is the deposition fraction in the ith airway, Ai the activity in the ith airway, Afs the activity on the filter sample, Qfs the filter sample flow rate, MV the animal minute volume, tfs the filter sampling time, and texp the exposure time.
Particle deposition fractions for the URT and lung of rats for head-only and whole-body exposures were calculated using equation (1). Activity measurements for the stomach, esophagus, and duodenum were included in the URT deposition fraction. Deposition fraction in the URT and lung increased with particle aerodynamic diameter and was smaller than that of Raabe, et al. (1977, 1988). Differences in the exposure chambers, and associated differences in particle inhalability, aerosol mixing, and losses in the chambers, and the influence of expired air on particle concentration may explain the discrepancy.
Measured lung and total deposition fractions for head-only exposures were larger than those for whole-body exposures. Differences in deposition fractions between two exposure methods were due to a greater particle mixing and a smaller particle inhalability in the whole-body exposure scenario. While particle deposition in whole-body exposures increased with particle size, deposition in the head-only exposure increased with particle size for particle diameters up to 3 mm and decreased thereafter. Since particle deposition is expected to increase with particle size, the decrease in deposition is due to reduction in particle inhalability with increasing particle size (inertia).
A significant portion of inhalation exposures are nose-only. Thus a followup study was conducted to measure deposition of radiolabeled monodisperse particles in the lungs of female Long-Evans rats following a short exposure to particles ranging in size from 0.9 to 4.2 mm in a Cannon nose-only exposure. A condensation monodisperse aerosol generator (CMAG) (Model 3475; TSI Inc., St. Paul, MN) was used for the generation of monodisperse aerosols. Long-Evans rats were loaded into nose-only tubes (Skornik Plethysmograph, CH Technologies Inc., Westwood, NJ) 10-15 minutes before exposure to aerosol. A total of 11 exposure events were conducted. Animal respiratory parameters were measured for a 5-minute period prior to aerosol exposure and continuously during exposure. Animals were killed immediately at the end of the exposures, and particle deposition in the head and lung were measured.
The deposition fractions in the left lung and caudal, medial, cranial, and accessory lobes of the right lung were found from deposition measurements. Deposition patterns were similar among the lobes of the lung. Particle deposition losses increased with particle diameter for a particle diameter smaller than 3.5 µm, and declined thereafter due to the filtering effect of the URT. This trend also was observed in the deposition measurements of Raabe, et al. (1988) in Fischer 344 rats (Menache, et al., 1995). The calculated deposition fraction in the exposed animals showed some scattering at a particle size due to differences in their lung geometry and physiology parameters. The left lung received higher deposition of particles than each lobe of the right lung. In the right lung, the caudal and accessory lobes had the highest and lowest deposition respectively. Depositions in the right medial and right cranial lobes were similar.
Head deposition fractions increased initially to reach a maximum at a particle size around 2.5 µm before declining. Most deposition occurred in nasal passages. The maximum lung deposition occurred for a particle diameter near 3.5 µm and most likely was due to inertial losses in the upper airways of the LRT. Due to a strong nasal filtering, the lung deposition fraction was less than 10 percent for most particle sizes. Considerable variation in deposition results was observed since each animal breathed differently and had a different lung deposition fraction for each particle size.
When particle size exceeds a certain value in a breathing scenario, inertia prevents the particle from entering the respiratory tract. Inhalability fraction (aspiration efficiency) can be calculated from comparison of deposition measurements with theoretical prediction of deposition at different sizes. Particle inhalability fraction depends on particle size, as well as animal breathing parameters, and is calculated from
(2)
where DF and Δ are the measured and theoretical (100 percent inhalable) lung regional or total deposition fractions. Values for Δ can be found from studies where 100 percent of particles are inhalable, or alternatively from experimentally validated mathematical models. Anjilvel and Asgharian (1995) introduced a mathematical model of particle deposition in the rat lung reconstructed from morphometric lung measurements of Long-Evans rats (Raabe, et al., 1976). The model was used here for Δ calculations using the measured breathing parameters of the animals in the study. A predictive model of inhalability fraction in Long-Evans rats in a functional form similar to that proposed by Menache, et al. (1995) was found by fitting a function to the data.
(3)
This equation has the right trend by approaching 1 for small particles and 0 for large particles.
References:
Anjilvel S, Asgharian B. A multiple-path model of particle deposition in the rat lung. Fundamental and Applied Toxicology 1995;28(1):41-50.
Menache MG, Miller FJ, Raabe OG. Particle inhalability curves for humans and small laboratory animals. American Occupational Hygiene 1995;39(3):317-328.
Raabe OG, Yeh HC, Schum GM, Phalen RF. Tracheobronchial geometry: human, dog, rat, hamster, Report LF-53. Lovelace Foundation, Albuquerque, NM, 1976.
Raabe OG, Yeh HC, Newton GJ, Phalen PF, Velasquez DJ. Deposition of inhaled monodisperse aerosols in small rodents. Walton, WH, ed. In: Inhaled Particles, IV. Pergamon Press, Oxford, United Kingdom, 1977.
Raabe OG, Al-Bayati MA, Teague SV, Rasolt A. Regional deposition of inhaled monodisperse coarse and fine aerosol particles in small laboratory animals. The Annals of Occupational Hygiene 1998;32:53-63.
Journal Articles on this Report : 4 Displayed | Download in RIS Format
Other project views: | All 7 publications | 4 publications in selected types | All 4 journal articles |
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Asgharian B, Kelly JT, Tewksbury EW. Respiratory deposition and inhalability of monodisperse aerosols in Long-Evan rats. Toxicological Sciences 2003;71(1):104-111. |
R827996 (Final) |
Exit |
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Kelly JT, Kimbell JS, Asgharian B. Deposition of fine and coarse aerosols in a rat nasal mold. Inhalation Toxicology 2001;13(7):577-588. |
R827996 (2000) R827996 (Final) |
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Kelly JT, Bobbitt CM, Asgharian B. In vivo measurement of fine and coarse aerosol deposition in the nasal airways of female Long-Evans rats. Toxicological Sciences 2001;64(2):253-258. |
R827996 (Final) |
Exit |
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Kelly JT, Tewksbury EW, Wong BA, Asgharian B. Nasal and lung deposition of fine and coarse particles in rats. Annals of Occupational Hygiene 2002;46(Supplement 1):346-349. |
R827996 (Final) |
not available |
Supplemental Keywords:
Long-Evans rats, deposition efficiency, deposition fraction, nasal airway, impaction, in vivo deposition, nasal mold, particle inhalability fraction, dose-response., RFA, Scientific Discipline, Air, particulate matter, Toxicology, air toxics, Environmental Chemistry, Health Risk Assessment, Atmospheric Sciences, ambient aerosol, urban air, inhalability, exposure and effects, dose response, air pollution, chronic health effects, particulate exposure, mortality, lower respiratory tract injury, animal inhalation study, dosimetryProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.